Evaluation of a single-plasmid PstDNV virus-like particle system for dsRNA delivery against WSSV in Litopenaeus vannamei

Information from the abstract

White Spot Syndrome Virus (WSSV) threatens shrimp aquaculture, causing rapid mortalities and severe economic losses. Shrimp rely on RNA interference (RNAi) as a sequence-specific antiviral defense; however, naked double-stranded RNA (dsRNA) can be unstable in shrimp hemolymph. Virus-like particles (VLPs) provide biocompatible protein shells that can improve dsRNA stability and delivery. In multi-plasmid co-expression systems, variable plasmid copy number can hinder the balance between dsRNA and capsid production, reducing dsRNA encapsulation. Here, a single-plasmid system was engineered in Escherichia coli to express the capsid protein of Penaeus stylirostris densovirus ( Pst DNV) and dsRNA targeting the WSSV ribonucleotide reductase subunit 2 ( rr2 ) gene, enabling in-cell auto-encapsulation of dsRNA into the VLPs. The cp Pst DNV-ds rr2 complex formed uniform VLPs (∼20–22 nm), verified by SDS-PAGE, Western blotting, LC-MS/MS, and transmission electron microscopy (TEM). Encapsulated dsRNA was protected from RNase III digestion and showed increased resistance to enzymatic degradation in shrimp hemolymph during the early exposure window, while VLP integrity was maintained during storage at 4 °C and −80 °C for up to six months. In a WSSV challenge bioassay, L. vannamei treated with cp Pst DNV-ds rr2 complexes had reduced relative WSSV DNA levels and improved survival by up to 60% compared to control shrimp that received NaCl only. Compared with naked dsRNA, the cp Pst DNV-ds rr2 complex showed lower protection, reflected by mortality rates that were 20% higher on day 1 and 10% higher on day 5. This trade-off suggests that encapsulation stabilizes dsRNA but may delay intracellular release, underscoring the need to optimize release kinetics and improve dsRNA yield and loading during bacterial production for aquaculture applications.

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Related topics: Invertebrate Immune Response Mechanisms · Viral Infectious Diseases and Gene Expression in Insects · Virus-based gene therapy research